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OLIGODENDROCYTES IN CNS DEVELOPMENT, DISEASE AND MYELIN REGENERATION David Rowitch, MD, PhD Departments of Pediatrics and Neurosurgery Howard Hughes Medical Institute UCSF

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Objectives <ul><li>Understand fundamental principles of oligodendrocyte development from neural stem cells. </li></ul><ul><li>Appreciate the rational for various therapeutic approaches that might alter the reparative functions of oligodendrocytes. </li></ul><ul><li>Understand basis for neural stem cell Phase I clinical trial for the congenital leukodystrophy, Pelizaeus-Merzbacher Disease, at UCSF. </li></ul>

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<ul><li>Demyelinating disease </li></ul><ul><li>Most common neurological disease </li></ul><ul><li>in young adults </li></ul><ul><li>Incidence ~1:1000; W>M </li></ul><ul><li>Pathogenesis: ?Autoimmune, acute; deficient </li></ul><ul><li>repair, sub-acute/chronic. </li></ul><ul><li>Congenital leukodystrophy </li></ul><ul><li>Incidence <1/500,000 </li></ul><ul><li>Total absence of myelin </li></ul>Pelizaeus-Merzbacher Disease (PMD) Multiple sclerosis (MS)

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Oligodendrocyte function in myelination Saltatory (jumping) nerve conduction

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Franklin and ffrench-Constant, 2008, Nat. Neurosci. Rev. Two outcomes of MS lesions Repair Chronic demyelination MS progression comprises (1) Autoimmune attack on OL, (2) inability to effectively repair in timely manner

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OLIG1/2 in CNS Development and Disease Lu et al., 2000, Neuron ; Zhou et al., 2000, Neuron ; Lu et al., 2002, Cell ; Zhou and Anderson, 2002; Cell; Takebayashi et al., 2002, Curr. Biol. ; Ligon, Huillard et al., 2007, Neuron; Arnett, Fancy et al., 2004, Science; Sun et al., 20111, Neuron ; Mehta et al., 2011, Cancer Cell Richard Lu (UTSW) Chuck Stiles (DFCI) Heather Arnett (Amgen) Steve Fancy (UCSF) Robin Franklin (Cambridge) Embryonic Neural Tube

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OLIG1 expression in multiple sclerosis Olig1 nuclear Olig1 cytoplasmic Unaffected white matter Edge MS plaque Arnett, Fancy et al., 2004, Science

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Demyelinated axons OLP Focal area of OL death: demyelination Myelinated tissue Lysolethicin gliotoxic injury model Remyelination Focal lysolecithin injection (Solochome cyanine)

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Olig1 is required for remyelination Focal lysolecithin injection (Solochome cyanine) Arnett, Fancy et al., 2004, Science Olig1 -/- Wild-type 14 dpl 28 dpl Ultra structure

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Developmental perspective Chronic demyelination “ Blocked” OLP are present in human MS lesions NG2 PDGFR  Pre-OL OLP OL Olig2 NG2 Olig1 (Nuclear) PLP Olig1 (Cytoplasmic) APC (CC1) O4 Nkx2.2 O1 APC Axin2 Are OLP blocked in MS?

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What Programs Regulate Myelin Repair?

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Genomic Screen: Remyelination kinetics Fancy et al., 2009, Genes Dev Solochrome cyanine Tissue array Genomic Expression Screen In situ hybridization with 1040 probes White matter lesions OL differentiation 5dpl Myelination OLP recruitment 10dpl 14dpl ~50 Transcription factor- encoding genes Transcription factor screen ~70% of TF genes in mammals ~50 genes with dynamic regulation during remyelination

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Tcf4 (and Wnt reporter) expressed during OLP development and remyelinating OLP Solochrome cyanine Tissue array Genomic Expression Screen In situ hybridization with 1040 probes White matter lesions Focal lysolecithin injection Focal ethidium bromide injection Dietary cuprizone Mouse SC Mouse CC Rat CCP OL differentiation 5dpl Myelination OLP recruitment 10dpl 14dpl ~50 Transcription factor- encoding genes Genomic Screen Identifies Tcf4 Tcf4 is a mediator of Wnt signaling Fancy et al., 2009, Genes Dev Tcf4 Olig2 Tcf4 Olig2 Tcf4 Olig2

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Tcf4 mediates Wnt-Catenin Signaling Inactive Activated Axin2

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OLP markers unchanged; OL differentiation delayed Axin2, Nkd1, Notum expression elevated P4 Olig2-cre X (fl)DA-Catenin vs. wild type spinal cord Affy mRNA array 430 2.0 Wnt-induced myelination delay

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Fancy et al., 2009, Genes Dev Wnt-induced remyelination delay WT Olig2cre X DA-  -Cat PLP Nkx2.2 10 dpl

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Model: Wnt signaling in oligodendrocytes Pre-OL OLP OL APC Axin2? Active Wnt-Catenin signaling inhibits the transition to mature oligodendrocyte

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<ul><li>Dual Role for Axin2 in remyelination? </li></ul><ul><li>Readout of endogenous Wnt activity </li></ul><ul><li>Inhibitor of pathway </li></ul>

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Delayed remyelination in Axin2 -/- mice

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Huang et al., 2009, Nature Axin2 required to inhibit Wnt pathway

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XAV939 stabilizes Axin2 in OLP in vitro Huang et al., 2009, Nature Fancy et al., Nature Neuroscience, In press

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Add XAV939 small molecule Wnt inhibitor… Wnt inhibitor accelerates remyelination Fancy et al., Nature Neuroscience, In press OLP differentiation and remyelination accelerated ~4 days (Axin2-/- mice unaffected by XAV)

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XAV939 does not affect inflammatory, gliotic response, OL death, OLP recruitment or proliferation Wnt inhibitor accelerates remyelination

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Pre-OL OLP OL APC Axin2 Developmental perspective PDGFR  Olig2 NG2 Olig1 (Nuclear) PLP Olig1 (Cytoplasmic) APC (CC1) O4 Nkx2.2 O1 MS

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NSC Replacement NSC Regulation Perspective on neural progenitor transplant versus regulation Pre-OL OLP OL MS

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Pelizaeus-Merzbacher Disease (PMD) <ul><li>X-linked recessive mutation of PLP1 </li></ul><ul><li>(Loss-of-function or gene duplications) </li></ul><ul><li>Fatal form: onset in infancy; affected males fail to achieve motor milestones; death by ~5 years. </li></ul><ul><li>Oligodendrocyte cell type-specific disease </li></ul>

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Normal (age 2 years) PMD (age 2 years) <ul><li>MRI T2-weighted images. </li></ul>Pelizaeus-Merzbacher Disease (PMD)

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Restricted origins, diffuse migration and transplant of oligodendrocytes Pringle et al., 1993 Neuron ; Orentas et al., 1999 Development ; Rowitch, 2004 Nat. Neurosci. Rev. ; Windrem et al., 2008 Cell Stem Cell Human OL in Shi mouse brain. Work of Goldman lab, U. Rochester.

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Phase I clinical trial for PMD <ul><li>Considerations: PMD as disease target </li></ul><ul><li>Lack of inflammation. </li></ul><ul><li>Conatal form of PMD is fatal </li></ul><ul><li>PMD patient ’s brains are profoundly hypomyelinated. </li></ul><ul><li>Myelin can be sensitively/non-invasively detected by MRI, </li></ul><ul><li>so myelination serves as definitive indicator of donor cell engraftment/function, without need to label cells. </li></ul><ul><li>StemCells, Inc. received FDA approval Dec. 2008. </li></ul><ul><li>Phase I Trial Jan. 2010  Feb. 2012 </li></ul><ul><li>Determine safety of human neural progenitor (StemCells, Inc.) transplant in patients with severe/fatal PMD. </li></ul><ul><li>Concerns: tolerance of cells, immunosuppressive therapy, tumor development. </li></ul>

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Summary <ul><li>Function for Olig genes. Utility of these and other markers in assessment of human WMI. </li></ul><ul><li>MS WMI involves failure of remyelination. Regulation of OLP repair by Wnt signaling </li></ul><ul><li>Consideration of stem cell therapy for fatal leukodystrophies </li></ul>

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XAV939

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Acknowledgements Steve Fancy Vivi Heine Emily Harrington Tracy Yuen John Silberis Jose Otero Eric Huang David Rowitch UCSF Richard Lu (UTSW) Heather Arnett (Amgen) Charles Stiles Dana-Farber Cancer Institute Chao Zhao Robin Franklin Cambridge University Saraid Billards Hannah Kenney Children ’s Hospital, Boston Stephen Huhn Stem Cells, Inc. Roel Nusse Stanford University